With the long term of understanding regulated organellar assembly, we propose to focus on the lysosome. The lysosome and lysosomal storage diseases are one of the clearest examples of human disease states from which hypotheses for the regulation of organelle assembly can be based. The morphological retention of lysosomes in I cell fibroblasts, cells from patients suffering from mucolipidosis II in which lysosomal hydrolases are secreted, indicates that lysosomal membrane synthesis and hydrolase synthesis are differentially regulated. We propose to investigate this hypothesis under two different regulatory conditions. In the first, there is an apparent "induction" of hydrolase accumulation. In the second, the amine treated or mutant fibroblast, there is extensive secretion of lysosomal hydrolases while cells still maintain abundant lysosomes. In these two cases, we will compare lysosomal membrane protein biosynthesis (and localization) with that of lysosomal hydrolases. In addition to these studies, we propose to determine the mechanism(s) by which newly synthesized lysosomal proteins are transmitted through the organelle population. To do this, experiments will be done to characterize lysosomal proteins, to establish the kinetics of their transfer from Golgi apparatus to lysosomes in vivo and the effects of drugs on this process, and to investigate the exchange of lysosomal molecules between lysosomes by organelle "fusion"/transfer processes. For experiments to establish molecule exchange within the lysosome population, cells with differentially labeled lysosomes will be fused either by viral infection or by transfection with a fusogen protein. Much of the methodology of contemporary cell biology including cell fractionation, antibodies and immunolocalization, cell fusion, radiolabeling, video microscopy, and gel electrophoresis will be employed in these studies.